【Description】

• TOROIVD^®5G qPCR Premix with UNG is a fast 2×master mix that provides for sensitive, reproducible detection up to five DNA targets. Particularly useful for virus detection with TaqMan^®probe assays, this premix includes TOROIVD^®5G DNA polymerase, dNTPs, dUTP, UNG and reaction buffer. The improved enzymes and reaction mixture combination also enables a high resistance to PCR inhibitors and high stability in room temperature. The premix is suitable for high-throughput analysis and can reduce the risk of cross-contamination with UNG. The premix is suitable for high-speed qPCR and enables accurate detection and quantification of targets, making it possible to obtain highly reproducible and reliable real-time PCR results over a wide dynamic range.

【Feature】

• -Rapid and highly sensitive

   This premix can achieve the rapid and highly sensitive quantification of a low-copy targets with probes and be suitable for the quantification of DNA viruses or cDNA at a low level.

• -Wide dynamic range

   This premix has been optimized to provide high specificity and dynamic range for use with DNA targets. This input flexibility can help streamline the number of different workflows in your lab to improve efficiency.

• -Optimized for multiplexing

  This premix has been validated for multiplexing up to five targets simultaneously, allowing for additional targets and/or controls to be run simultaneously for efficiency or quality control purposes.

• -Avoid Contamination

  This premix contains dUTP  and UNG in the reaction buffer. The crossed contamination caused by PCR product can be removed so that the rate of false-positive detection can be reduced.    

• -Room-temperature stable: 

  The specially optimized PCR buffer make the mix very stable at room temperature. Therefore, the performance is not easily decrease during storing and shipping.

【Components】

•  QPT-200U can be used for 500 reactions for a total 20µL reaction volume or 400 reactions for a total 25µL reaction volume.

• Cat NO.	Components	Size
  QPT-200U	TOROIVD®5G qPCR Premix with UNG	1.25 mL ×4 tubes/ bag

  
  

  
  

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• Notes: 2×5G qPCR Premix with UNG contains 0.4mM dA/C/G/T/UTP, 5mM Mg2+, UNG, TOROIVD^®5G DNA polymerase, reaction buffer and stabilizer，etc.

【Storage】

• This reagent can be stored at 2-8°C for 12 months. 

• For longer storage, this reagent should be kept at -20°C. 

【Primer/Probe Design】

• -Design of primers

  Primer length: 18–25bp；Tm of primer: 60–65°C；GC content: 40–60%；Target length: 70–200 bp;

  Larger targets (>200 bp) tend to reduce the efficiency and specificity of amplification.

  Purification grade: OPC or HPLC grade;

• -Design of probes

  Probe length: 20–30bp; Tm of probe: 65–70°C; GC content: 40–60%; Purification grade: HPLC.

• -Design for avoiding Contamination

  Amplicon with high GC content will degrade the the performance of UNG for removing the contamination caused by PCR product. Therefore, when designing the primers, it is necessary to minimize the GC content of the amplicon as much as possible.

• -Checking the performance of primers and probes：

  Prepare a dilution series with five or more dilutions of template DNA. Perform qPCR assay using the diluted DNA with the newly designed primers and probe, and draw a standard curve. Confirm that the PCR efficiency is between 90% and 110% and R2 is equal to or greater than 0.99. If the PCR efficiency or R2 are outside of these ranges, the reaction conditions should be optimized. If this does not improve the result, the primers and/or probe should be redesigned. 

【Application Data】

  1. High sensitivity verification

      -Template DNA：a plasmid with the B646L gene of African swine fever virus (VP72 protein) 

      -Primer&Probe sets： From WOAH.

• Conclusion: QPT-200U can stably detect ASFV in 3-4 copies/reaction, with high PCR efficiency and sensitivity.

  2.Thermal Stability Verification

     -Template DNA: a plasmid with the B646L gene of African swine fever virus (VP72 protein) 

     -Primer&Probe sets: From WOAH.

• -Reagents: QPT-200U stored at -20 ℃ and 37 ℃ for  30 days.

   

• Conclusion: There is no significant difference in the performance of QPT-200U after 30 days  at 37 ℃ and  at -20 ℃. QPT-200U can be stored at 2-8 ℃ and shipped at room temperature within 30 days. It reflects extremely strong thermal stability, reduces shipping costs.

【Documents】

•     Manuals:   TOROIVD®5G qPCR Premix with UNG (QPT-200U)- English-

•     Brochures:   Real-time PCR Premix for gene expression analysis

•     MSDS： Material Safety Data Sheet for QPT-200U

•     COA： Certificate of Analysis for QPT-200U  

    

【Order information】  

• Bulk package of 100mL can be supplied，please contact us .